Rapid and Direct Quantification of Viable Candida Species in Whole Blood by Use of Immunomagnetic Separation and Solid-Phase Cytometry

Abstract

Candida species are a common source of nosocomial bloodstream infections in critically ill patients. The sensitivity of the traditional diagnostic procedure based on blood culture is variable, and it usually takes 2 to 4 days before growth of Candida species is detected. We developed a 4-h method for the quantification of Candida species in blood, combining immunomagnetic separation (IMS) with solid-phase cytometry (SPC) using viability labeling. Additionally, Candida albicans cells could be identified in real time by using fluorescent in situ hybridization. By analysis of spiked blood samples, our method was shown to be sensitive and specific, with a low detection limit (1 cell/ml of blood). In a proof-of-concept study, we applied the IMS/SPC method to 16 clinical samples and compared it to traditional blood culture. Our method proved more sensitive than culture (seven samples were positive with IMS/SPC but negative with blood culture), and identification results were in agreement. The IMS/SPC data also suggest that mixed infections might occur frequently, as C. albicans and at least one other Candida species were found in five samples. Additionally, in two cases, high numbers of cells (175 to 480 cells/ml of blood) were associated with an endovascular source of infection.

Candida glabrata: Pathogenicity and Resistance Mechanisms for Adaptation and Survival

“…Major risk factors for Candida infections include prolonged usage of broad-spectrum antibiotics, immunocompromised state of the host, and the use of medical devices in surgery including catheters [ 3 , 4 ]. Candida species commonly cause invasive nosocomial infections in immunocompromised patients [ 5 ]. It accounts for 70–90% of all aggressive mycoses [ 6 ].…”

Rapid identification of yeast by fluorescence in situ hybridisation from broth and blood cultures

“…Fluorescence signals from different species‐specific FISH probes might occur in case of mixed bloodstream infections with different Candida spp. The occasional occurrence of such mixed infections was demonstrated by immunomagnetic separation and solid‐phase cytometry 58 . Mixed infections might be missed by conventional biochemical differentiation methods or MALDI‐TOF MS if only a single colony is chosen for further differentiation from a non‐selective agar, as is often done in routine laboratory procedure, in particular in cases of morphologically indistinguishable colonies.…”

Fungal infections diagnosis – Past, present and future

“…Until the final result of the microorganism identification, the sample goes through a series of steps [ 154 ]. The sample is first filtered on a membrane and then retained cells are fluorescently labelled.…”

“…Fluorescent cells are analysed using a solid-phase cytometer, where background signals are distinguished from specific signals referring to target cells. Finally, the sample is analysed using fluorescence microscopy, in order to validate and examine the target cells [ 153 , 154 ].…”